The overall objective of this research is to clearly resolve the question of the number and affinities of ligand binding sites on individual mammalian IgM antibodies. The hypothesis to be tested is that the heterogeneity of ligand binding affinities exhibited by some IgM antibodies is an intramolecular phenomenon. Furthermore it is proposed that these intramolecular ligand binding differences may be due to intramolecular conformational differences resulting from differing modes of intracellular assembly and/or carbohydrate addition and not from primary structural difference. The specific aims to be accomplished are: a) to develop two different types of murine hybridomas secreting large amounts of IgM antibodies to defined ligands, i.e. one form secreting IgM molecules with five high and five low affinity ligand binding sites, b) to establish that the above mentioned high and low affinity sites represent an intramolecular phenomenon and are localized to different Fab fragments having the same primary structure, and c) to establish the molecular and/or cellular basis for the binding differences between the high and low affinity sites. Successful completion of this work will not only answer a long standing question regarding the structure and function of IgM, but will also likely raise important additional questions on the functional role(s) of IgM antigen receptors on B lymphocytes.